Two partially inbred strains of the laboratory opossum, Monodelphis domestica, exhibit extreme differences in plasma very low density and low density lipoprotein cholesterol after they are fed a high cholesterol, high fat diet; they exhibit somewhat less extreme differences in response to a diet enriched in cholesterol alone. Plasma cholesterol levels of high and low responding opossums are similar when they are maintained on a basal diet with moderate levels of cholesterol and fat, or when they are challenged with a diet enriched with fat alone. Pedigree data indicate that a recessive gene is largely responsible for the high levels of plasma cholesterol in high responding opossums. The overall goal of this project is to identify differences in the gene expression patterns of high and low responders in response to dietary cholesterol in order to advance our understanding of the genetic factors and physiological mechanisms that control lipemic responsiveness to dietary cholesterol. Fulfillment of this goal may enable the identification of the recessive gene responsible for diet-induced hypercholesterolemia in the opossum model. Suppression subtractive hybridization will be used to enrich differentially expressed genes in livers and small intestines of high and low responders fed a high cholesterol diet. These differentially expressed genes will be cloned and sequenced to reveal their identities. Some of these genes with known or potential biological significance in cholesterol and bile acid metabolism will be analyzed for sequence variations between the two strains in order to identify candidate genes that may be the primary gene responsible for the inter-strain difference. A genetic analysis of F2 progeny will be conducted to determine if any of the candidate genes is linked to the quantitative trait locus that is directly responsible for controlling dietary responsiveness. The results of these proposed studies will lead to new opportunities to exploit this unique model in experimental investigations of dietary responsiveness and cholesterol metabolism, and in developing and testing novel drugs for preventing hypercholesterolemia in humans.